The transplantation of organs is now practicable, due to major advances in surgical and other techniques. However, availability of suitable human organs for transplantation is a significant problem. Demand outstrips supply. This has caused researchers to investigate the possibility of using non-human organs for transplantation.
Xenotransplantation is the transplantation of organs from one species to a recipient of a different species. Rejection of the transplant in such cases is a particular problem, especially where the donor species is more distantly related, such as donor organs from pigs and sheep to human recipients. Vascular organs present a special difficulty because of hyperacute rejection (HAR).
HAR occurs when the complement cascade in the recipient is initiated by binding of antibodies to donor endothelial cells.
Previous attempts to prevent HAR have focused on two strategies: modifying the immune system of the host by inhibition of systemic complement formation (1,2), and antibody depletion (3,4). Both strategies have been shown to prolong xenograft survival temporarily. However, these methodologies are therapeutically unattractive in that they are clinically impractical, and would require chronic immunosuppressive treatments. Therefore, recent efforts to inhibit HAR have focused on genetically modifying the donor xenograft. One such strategy has been to achieve high-level expression of species-restricted human complement inhibitory proteins in vascularized pig organs via transgenic engineering (5-7). This strategy has proven to be useful in that it has resulted in the prolonged survival of porcine tissues following antibody and serum challenge (5,6). Although increased survival of the transgenic tissues was observed, long-term graft survival was not achieved (6). As observed in these experiments and also with systemic complement depletion, organ failure appears to be related to an acute antibody-dependent vasculitis (1,5).
In addition to strategies aimed at blocking complement activation on the vascular endothelial cell surface of the xenograft, recent attention has focused on identification of the predominant xenogeneic epitope recognised by high-titre human natural antibodies. It is now accepted that the terminal galactosyl residue, Gal-α(1,3)-Gal, is the dominant xenogeneic epitope (8-15). This epitope is absent in Old World primates and humans because the α(1,3)-galactosyltransferase (gal-transferase or GT) is non-functional in these species. DNA sequence comparison of the human gene to α(1,3)-galactosyltransferase genes from the mouse (16,17), ox (18), and pig (12) revealed that the human gene contained two frameshift mutations, resulting in a nonfunctional pseudogene (20,21). Consequently, humans and Old World primates have pre-existing high-titre antibodies directed at this Gal-α(1,3)-Gal moiety as the dominant xenogeneic epitope.
One strategy developed was effective to stably reduce the expression of the predominant Gal-α(1,3)-Gal epitope. This strategy took advantage of an intracellular competition between the gal-transferase and α(1,2)-fucosyltransferase (H-transferase) for a common acceptor substrate. The gal-transferase catalyzes the transfer of a terminal galactose moiety to an N-acetyl lactosamine acceptor substrate, resulting in the formation of the terminal Gal-α(1,3)-Gal epitope. Conversely, H-transferase catalyzes the transfer of a fucosyl residue to the N-acetyl lactosamine acceptor substrate, and generates a fucosylated N-acetyl lactosamine (H-antigen, i.e., the O blood group antigen), a glycosidic structure that is universally tolerated. Although it was reported that expression of human H-transferase transfected cells resulted in high level expression of the non-antigenic H-epitope and significantly reduced the expression of the Gal-α(1,3)-Gal xenoepitope, there are still significant levels of Gal-α(1,3)-Gal epitope present on such cells.